The number of prokaryotes and the total amount of their cellular carbon on earth are estimated to be 4-6 3 10 30 cells and 350-550 Pg of C (1 Pg 5 10 15 g), respectively. Thus, the total amount of prokaryotic carbon is 60-100% of the estimated total carbon in plants, and inclusion of prokaryotic carbon in global models will almost double estimates of the amount of carbon stored in living organisms. In addition, the earth's prokaryotes contain 85-130 Pg of N and 9-14 Pg of P, or about 10-fold more of these nutrients than do plants, and represent the largest pool of these nutrients in living organisms. Most of the earth's prokaryotes occur in the open ocean, in soil, and in oceanic and terrestrial subsurfaces, where the numbers of cells are 1.2 3 10 29 , 2.6 3 10 29 , 3.5 3 10 30 , and 0.25-2.5 3 10 30 , respectively. The numbers of het- erotrophic prokaryotes in the upper 200 m of the open ocean, the ocean below 200 m, and soil are consistent with average turnover times of 6-25 days, 0.8 yr, and 2.5 yr, respectively. Although subject to a great deal of uncertainty, the estimate for the average turnover time of prokaryotes in the subsurface is on the order of 1-2 3 10 3 yr. The cellular production rate for all prokaryotes on earth is estimated at 1.7 3 10 30 cellsyyr and is highest in the open ocean. The large population size and rapid growth of prokaryotes provides an enormous capacity for genetic diversity. Although invisible to the naked eye, prokaryotes are an essential component of the earth's biota. They catalyze unique and indispensable transformations in the biogeochemical cy- cles of the biosphere, produce important components of the earth's atmosphere, and represent a large portion of life's genetic diversity. Although the abundance of prokaryotes has been estimated indirectly (1, 2), the actual number of pro- karyotes and the total amount of their cellular carbon on earth have never been directly assessed. Presumably, prokaryotes' very ubiquity has discouraged investigators, because an esti- mation of the number of prokaryotes would seem to require endless cataloging of numerous habitats. To estimate the number and total carbon of prokaryotes on earth, several representative habitats were first examined. This analysis indicated that most of the prokaryotes reside in three large habitats: seawater, soil, and the sedimentysoil subsur- face. Although many other habitats contain dense populations, their numerical contribution to the total number of pro- karyotes is small. Thus, evaluating the total number and total carbon of prokaryotes on earth becomes a solvable problem. Aquatic Environments. Numerous estimates of cell density, volume, and carbon indicate that prokaryotes are ubiquitous in marine and fresh water (e.g., 3-5). Although a large range of cellular densities has been reported (10 4 -10 7 cellsyml), the

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Prokaryotes: The unseen majority

A comprehensive report of the acetylene reduction assay for measurement of N2 fixation is presented. The objective is to facilitate the effective use and identify some potential limitations of the method. The report is based on more than 200 accounts of the use of this technique in 15 countries during the last 5 years. Methods of measurement of N2 fixation are compared. Nomenclature, e.g., N2[C2H2] fixed, is introduced to identify values of N2 fixation determined by C2H2-C2H2 assay. The biochemical basis of the assay is described along with relevant characteristics including Km, C2H2/N2 conversion factor, and specific N2[C2H2]-fixing activities obtained with various systems. Effects of combined nitrogen, temperature, light, pO2, N2, pC2H2 and water on activity are summarized. Available methods for sample preparation, assay chamber, gas phase, assay condition, termination of reaction, C2H4 analysis and expression of results are compared. The many uses of the C2H2-C2H4 assay for investigations of the biochemistry of nitrogenase and physiology of N2-fixing organisms, definition of N2-fixing organisms and measurement of field N2 fixation by legume, non-legume, soil, marine, rhizosphere, phylloplane and mammalian samples are tabulated.

Applications of the acetylene-ethylene assay for measurement of nitrogen fixation

The extremely large number of leaves produced by terrestrial and aquatic plants provide habitats for colonization by a diversity of microorganisms. This review focuses on the bacterial component of leaf microbial communities, with emphasis on Pseudomonas syringae—a species that participates in leaf ecosystems as a pathogen, ice nucleus, and epiphyte. Among the diversity of bacteria that colonize leaves, none has received wider attention than P. syringae, as it gained notoriety for being the first recombinant organism (Ice− P. syringae) to be deliberately introduced into the environment. We focus on P. syringae to illustrate the attractiveness and somewhat unique opportunities provided by leaf ecosystems for addressing fundamental questions of microbial population dynamics and mechanisms of plant-bacterium interactions. Leaf ecosystems are dynamic and ephemeral. The physical environment surrounding phyllosphere microbes changes continuously with daily cycles in temperature, radiation, relative humidity, wind velocity, and leaf wetness. Slightly longer-term changes occur as weather systems pass. Seasonal climatic changes impose still a longer cycle. The physical and physiological characteristics of leaves change as they expand, mature, and senesce and as host phenology changes. Many of these factors influence the development of populations of P. syringae upon populations of leaves. P. syringae was first studied for its ability to cause disease on plants. However, disease causation is but one aspect of its life strategy. The bacterium can be found in association with healthy leaves, growing and surviving for many generations on the surfaces of leaves as an epiphyte. A number of genes and traits have been identified that contribute to the fitness of P. syringae in the phyllosphere. While still in their infancy, such research efforts demonstrate that the P. syringae-leaf ecosystem is a particularly attractive system with which to bridge the gap between what is known about the molecular biology of genes linked to pathogenicity and the ecology and epidemiology of associated diseases as they occur in natural settings, the field.

Bacteria in the leaf ecosystem with emphasis on Pseudomonas syringae-a pathogen, ice nucleus, and epiphyte.

Biofilms are a form of collective life with emergent properties that confer many advantages on their inhabitants, and they represent a much higher level of organization than single cells do. However, to date, no global analysis on biofilm abundance exists. We offer a critical discussion of the definition of biofilms and compile current estimates of global cell numbers in major microbial habitats, mindful of the associated uncertainty. Most bacteria and archaea on Earth (1.2 × 1030 cells) exist in the ‘big five’ habitats: deep oceanic subsurface (4 × 1029), upper oceanic sediment (5 × 1028), deep continental subsurface (3 × 1029), soil (3 × 1029) and oceans (1 × 1029). The remaining habitats, including groundwater, the atmosphere, the ocean surface microlayer, humans, animals and the phyllosphere, account for fewer cells by orders of magnitude. Biofilms dominate in all habitats on the surface of the Earth, except in the oceans, accounting for ~80% of bacterial and archaeal cells. In the deep subsurface, however, they cannot always be distinguished from single sessile cells; we estimate that 20–80% of cells in the subsurface exist as biofilms. Hence, overall, 40–80% of cells on Earth reside in biofilms. We conclude that biofilms drive all biogeochemical processes and represent the main way of active bacterial and archaeal life. In this Analysis article, Flemming and Wuertz calculate the total number of bacteria and archaea on Earth and estimate the fraction that lives in biofilms. They propose that biofilms are the most prominent and influential type of microbial life.

Bacteria and archaea on Earth and their abundance in biofilms

▪ Abstract The vast surface of the plant axis, stretching from root tips occasionally buried deeply in anoxic sediment, to apical meristems held far aloft, provides an extraordinarily diverse habitat for microorganisms. Each zone has to a greater or lesser extent its own cohort of microorganisms, in aggregate comprising representatives from all three primary domains of life—Bacteria, Archaea, and Eucarya. While the plant sets the stage for its microbial inhabitants, they, in turn, have established varied relationships with their large partner. These associations range from relatively inconsequential (transient epiphytic saprophytes) to substantial (epiphytic commensals, mutualistic symbionts, endophytes, or pathogens). Through recent technological breakthroughs, a much better perspective is beginning to emerge on the nature of these relationships, but still relatively little is known about the role of epiphytic microbial associations in the life of the plant.

The Ecology and Biogeography of Microorganisms on Plant Surfaces.

The phyllosphere: I. An ecologically neglected milieu

SummaryMeasurements of the amounts of anthrone- and ninhydrin-positive substances occurring in rain-water and dew on plants in Surinam have been made, as well as of the possible nitrogen gains and losses in the dew.Nitrogen fixation in detached leaves in association with an autochthonous phyllosphere population and in those enriched withAzotobacter sp.,Beijerinckia sp., orPseudomonas sp. are compared.Dry weight and total nitrogen increases of single leaves, or part of leaves, of Coffea, Gossypium, and Phaseolus floated on a nitrogen-free medium in petri dishes were determined at intervals of a few days and compared with a control at the start of the experiment.Gains in total nitrogen amounting to 20 to 105 per cent over the control were measured within two weeks. The increases were found in the leaves as well as in the culture medium and were dependent on the age of the leaf, on the light, and on the temperature. The energy substrates for bacterial nitrogen fixation were obviously furnished by the leaf, which increased in size and up to 200 per cent in dry weight.In the limited space of the petri dish the cultural conditions for active nitrogen fixation quickly deteriorated by the accumulation of metabolic products from both the leaf and the microvegetation. Heterotrophs and predatory protozoa eventually dominated the initial population. Earlier gains were then partly lost.The consequences of the biocoenosis of leaves and microbes for the vegetation are discussed.

The phyllosphere: III. Nitrogen fixation in the phyllosphere

SummaryIn connection with the as yet unexplained gains in nitrogen in young grassland, the phyllosphere of grasses, in particularTripsacum laxum Nash, has been studied on the experimental farm and in the laboratories of the ORSTOM (Organisation de Recherches Scientifiques Tropicales Outre-Mer) at Adiopodoumé (Ivory Coast, West Africa). Considerable concentrations of sugars were observed in rain water and dew (14–200 ppm) and in the stem run-off (up to 465 ppm). The values were exceeded by those of the moisture in the leaf sheaths (up to 104 ppm), which thus form a special milieu for nitrogen-fixing elements. The composition of the microbial population depends on the physical state of the leaves concerned,viz age, climate, and weather conditions. The population consists mainly of Gram-negative rods, yeasts, and coryneform bacteria. The possible significance of the nitrogen fixation in grassland is discussed.The investigation is being continued in the Microbiological Laboratory at Wageningen.

Jakoba Ruinen

Papers

The phyllosphere: I. An ecologically neglected milieu

The phyllosphere: III. Nitrogen fixation in the phyllosphere

The phyllosphere: V. The grass sheath, a habitat for nitrogen-fixing micro-organisms